Thin Layer Chromatography of Inkdyes

To perform Thin Layer Chromatography of ink/dyes
INTRODUCTION: Thin Layer Chromatography (TLC)
Thin-layer chromatography is a chromatography technique used to isolate non-

volatile mixtures that separates pigments, identifying molecules. While it has many
applications in a wide variety of industries, it is a particularly important technique
used in forensic labs, helping scientists determine if two pieces of text were written
by the same ink, which can often be an indicator of fraud.
Thin layer chromatography can be used to monitor the progress of a reaction,

identify compounds present in a given mixture, and determine the purity of a
substance. A number of enhancements can be made to the original method to
automate the different steps, to increase the resolution achieved with TLC and to
allow more accurate quantitative analysis. This method is referred to as "high-
performance TLC". On completion of the separation, each component appears as
spots separated vertically. Each spot has a retention factor (Rf) expressed as:
Rf = dist. travelled by sample / dist. travelled by solvent
AIM:
Our goal is to examine and to compare the quality of the results obtained by thin-layer
chromatography, high-performance thin layer chromatography test in the analysis of
samples of ink/dyes.
Thin layer chromatography as a qualitative method has shown good quality of results
that can be used for a good life prediction of the ammunition.To study how to perform
Thin Layer Chromatography of ink/dyes.
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PRINCIPLE:
Like other chromatographic techniques, thin-layer chromatography (TLC) depends on

the separation principle. The separation relies on the relative affinity of compounds
towards both the phases. The compounds in the mobile phase move over the surface
of the stationary phase. The movement occurs in such a way that the compounds
which have a higher affinity to the stationary phase move slowly while the other
compounds travel fast. Therefore, the separation of the mixture is attained. On
completion of the separation process, the individual components from the mixture
appear as spots at respective levels on the plates. Their character and nature are
identified by suitable detection techniques.
REQUIREMENT:
Chromatography jar Forceps (tweezers)

Micro-capillary tubes Methanol in dropper bottle
Mobile phase (solvent): 7.0 ml Ethyl
Ink sample Stirrer Beaker actetate + 3.5 ml ethanol + 3ml distilled
water
Glass slides or TLC plates Silica Gel G and
Ruler Pencil Scissors
Distilled water Hot air oven
PROCEDURE:
Thin Layer Chromatography Procedure

Before starting with the Thin Layer Chromatography Experiment, the different
components required to conduct the procedure along with the phases involved.
1. Obtain a silica gel plate (TLC paper), handle it by the edges to avoid
contaminating.
2. Draw a pencil line all the way

across the bottom of paper 2-3
mm up from the bottom.
3. If working with an ink source: mark
TLC paper at the starting line.
Note using a pencil the identity of
the sample being tested. More
than one sample can be tested on
the TLC paper.
2
If working with an ink on paper. First extract the ink sample from the paper.
Using a capillary tube, wick up an ink sample. Apply capillary tube to starting
line on the TLC paper to create a spot. Duly note on the TLC paper which
sample is applied. If working with dilute samples we can achieve better results
by drying the application and re-applying. Repeat until applied 3-5 times.
4. The TLC plate is now ready to place into a jar already adequately filled with
solvent (ethanol). The TLC plate will be positioned so that the bottom is in the
solvent but no solvent will be above the starting line we have marked with our
pencil. The solvent jar will be covered while the TLC plate is being developed in
order for the solvent vapors to equilibrate.
5. Allow the solvent to continue until all possible separations have occurred.
Typically the solvent is 80% of the way up the paper.
6. Remove the plate from the jar. Immediately mark the solvent front and the dye
spots with a pencil. Note all observations in our lab notebook including a
complete drawing in our notebook of the finished TLC plate. Measure and
record the distance each spot (from its center) moved from the starting line.
Calculate the Rf values for each. Some ink will have different components, and
thus will have multiple Rf values. Put Rf values in a table format to report.
7. Sometimes a critical component of the ink has fluorescence. Using an
ultraviolet light source we can observe our samples for this unique property.
RESULT:
As the solvent slowly travels up the plate,

the different components of the dye mixture
travel at different rates and the mixture is
separated into different coloured spots.
Measuring Rf values
Measurements are often taken from the plate in order to help identify the
compounds present. These measurements are the distance travelled by the
solvent, and the distance travelled by individual spots.
When the solvent front gets close to the top of the plate, the plate is removed from
the beaker and the position of the solvent is marked with another line before it has
a chance to evaporate.
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These measurements are then taken:
The Rf value for each dye is then worked out using the formula:
For example, if the red component travelled 1.7 cm from the base line while the
solvent had travelled 5.0 cm, then the Rf value for the red dye is:
If we could repeat this experiment under exactly the same conditions, then the Rf
values for each dye would always be the same. For example, the Rf value for the
red dye would always be 0.34. However, if anything changes (the temperature, the
exact composition of the solvent, and so on), that is no longer true. We have to
bear this in mind if we want to use this technique to identify a particular dye. We'll
look at how we can use thin layer chromatography for analysis further down the
page.
CONCLUSION:
TLC is one of the fastest, least expensive, simplest and easiest chromatography
technique. The technique of chromatography was first used in 1900 by scientist
Mikhail Tsvet to separate the pigments of plants. Since its invention, TLC has
evolved over the decades for use in numerous applications. One of the most
prominent applications of the technology is the separation of multicomponent
pharmaceutical formulations.
It is also heavily relied on for separating and identifying various colors,

preservatives, cosmetic products, and sweetening agents in the food and cosmetic
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sectors. In addition, forensic science has also adopted TLC that it is used in ink
analysis, usually helping to determine if a document has been forged.
Ink analysis is an incredibly important analytical technique in forensic crime labs.

Most commonly, it is used to determine if more than one ink was used on a
document, which can help to detect fraud and forgery which can provide invaluable
clues in criminal cases.

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To perform Thin Layer Chromatography of ink/dyes

INTRODUCTION: Thin Layer Chromatography (TLC)

Thin-layer chromatography is a chromatography technique used to isolate non-

Thin layer chromatography can be used to monitor the progress of a reaction,

Rf = dist. travelled by sample / dist. travelled by solvent

Like other chromatographic techniques, thin-layer chromatography (TLC) depends on

Chromatography jar Forceps (tweezers)

Thin Layer Chromatography Procedure

2. Draw a pencil line all the way

As the solvent slowly travels up the plate,

It is also heavily relied on for separating and identifying various colors,

Ink analysis is an incredibly important analytical technique in forensic crime labs.

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